The Click-iT™ EdU Alexa Fluor™ 555 HCS Assay is a superior alternative to traditional proliferation assays that is optimized for high-content imaging applications. In this assay the modified thymidine analogue EdU is efficiently incorporated into newly synthesized DNA and fluorescently labeled with a bright, photostable Alexa Fluor™ dye in a fast, highly-specific click reaction. This fluorescent labeling of proliferating cells is accurate and compatible with antibody methods due to the mild click protocol.

The Click-iT™ EdU Alexa Fluor™ 555 HCS Assay is:

• Simple—works the first time, every time, in less time
• Efficient—no denaturation steps or harsh treatment required
• Gentle on samples—better preservation of cell morphology, antigen structure, and DNA integrity
• Consistent—not dependent on variable antibody lots for detection

The most accurate proliferation-detection methods are based on the incorporation and measurement of nucleoside analogues in newly synthesized DNA, with bromodeoxyuridine (BrdU) a commonly used analogue. BrdU-labeled DNA is quantitated using anti-BrdU antibodies following DNA denaturation by harsh methods (HCl, heat, or enzymes) to expose the BrdU molecules. This step is time consuming and difficult to perform consistently. The harsh treatment can also adversely affect sample integrity and quality, which makes co-staining with other antibodies challenging.

Superior Proliferation Methodology
The Click-iT™ EdU Alexa Fluor™ 555 HCS Assay provides a superior alternative to BrdU assays for measuring cell proliferation. EdU (5-ethynyl-2'-deoxyuridine) is a nucleoside analog of thymidine and is incorporated into DNA during active DNA synthesis. With Click-iT™ EdU, mild fixation and detergent permeabilization is sufficient for the small molecule-based Click-iT™ EdU detection reagent to gain access to the DNA. As a consequence, the Click-iT™ EdU HCS assay is not only easy to use, but more accurate and compatible with cell cycle analysis and other intracellular or extracellular targets for truly content-rich results.

The kit is optimized for high-content imaging applications; visit the Click-iT™ technology area of our website for kits designed for fluorescence microscopy, microplate, or flow cytometry platforms.

Learn more about Click-iT™ technology

Notes:
The Click-iT™ assay can be used on cells in culture or in vivo following administration of EdU by feeding or injection methods.
The Click-iT™ assay can be used with BrdU in dual pulse experiments by using the ant-BrdU (clone MoBu-1) antibody, which does not cross react with EdU.
The Click-iT™ technology is compatible with immunohistochemical, immunocytochemical, and fluorescent dyes that are fixation tolerant or designed for fixed-cell labeling.